Several different sources of cells for the generation of insulin producing cells beta-cells for the treatment of diabetes can be envisioned. These include pluripotent stem cells as well as adult endoderm derivatives such as liver cells. The current focus in producing beta-cells from pluripotent precursors is on the use of extrinsic factors, whereas the generation of beta-cell from hepatic cells involves genetic reprogramming using gene transfer strategies. This project brings together investigators working on these different approaches for the purpose of integrating relevant information and using it to guide us towards the most efficient way of producing transplantable beta-cell equivalents. Specifically, epigenetic analysis of differentiation intermediates from all projects will be used to inform rational decisions about the extrinsic/genetic manipulations required in each system to achieve the final goal of therapeutically useful cells. Because in vivo reprogramming of liver cells requires the use of adenoviral vectors, the role of this virus in reprogramming must be elucidated. Heterogeneity of the in vitro products is expected in each system and hence reagents to purify more epigenetically homogeneous populations are needed. We propose further development of surface reactive monoclonal antibodies for this purpose. Finally, functional validation of the differentiation products requires in vivo testing upon transplantation into animal models. PUBLIC HEALTH RELEVANCE: Transplantation of islets from cadaveric donors has shown that type 1 diabetes can be successfully treated by cell therapy. However, high quality cadaveric islet donors are rare and new sources of transplantable beta-cells must be found in order for this approach to realize its clinical potential. This proejct will produce patient-matched autologous beta-cells for the treatment of type 1 diabetes. Successful execution will impact large numbers of patients world-wide.